On-spot hydrophilic enhanced slide and preparation thereof

ABSTRACT

The invention discloses an on-spot hydrophilic enhanced slide/microarray. The preparation method relates to a hydrophobic copolymer prepared by blending, grafting or co-polymerization of a hydrophobic material and a compound bearing functional groups such as anhydride, imide, cyclic amide, and cyclic ester, and application of the hydrophobic copolymer onto an organic or inorganic substrate. The resulting slide has the properties of on-spot hydrophilic/hydrophobic dynamic conversion, as well as on-spot hydrophilic enhancement for the preparation of high-density and high-efficiency bio-chip/microarray.

CROSS-REFERENCE TO RELATED APPLICATION

The present application is a divisional application of U.S. patentapplication Ser. No. 09/695,254 filed on Oct. 25, 2000, now issued asU.S. Pat. No. 6,403,368, contents of which are incorporated herein byreference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to an on-spot hydrophilic enhanced slideand the preparation method thereof. More particularly, it relates to ahydrophobic copolymer prepared by blending, grafting orco-polymerization of a hydrophobic material and a compound bearingfunctional groups such as anhydride, imide, cyclic amide, and cyclicester, and application of the hydrophobic copolymer onto an organic orinorganic matrix to form an on-spot hydrophilic enhanced slide.

2. Description of the Related Arts

In the current bio-chip and bio-microarray technology, most of thepreparation methods involve the treatment of a matrix surface withsilanization, followed by the crosslinking reaction with biomaterials.In the silanization treatment, the surface of the substrate is activatedbased on its material, and then treated by a hydrophilic silane such asAPTES. Afterwards, the crosslinking reaction is performed via acrosslinker such as glutaldehyde to immobilize biomaterials on thesubstrate. The shortcomings of the method include substrate dependence,long reaction time, poor homogeneity, low reaction efficiency, and theresulting low activity for the immobilized biomaterials. Moreover, theprepared covalent bonding surface is hydrophilic, which facilitates thephenomenons of crossover and contamination among spots when thehydrophilic surface is used for a high-density microarray.

U.S. Pat. No. 5,837,860 and WO 98/39481 disclose the treatment of glassor silicon wafer with hydrophobic silane such as mercapto-silane, andthe immobilization of nucleic acid probes thereon. The method involvestreating a substrate surface so that mercapto-groups (HS-) withhydrophobicity are covalently bonded thereon. The hydrophobic propertyis suitable for the immobilization of nucleic acids/nucleotides in highdensity. The method, however, requires the modification of thebiomaterials to bear mercapto-groups, thereby forming disulfide bondsbetween the modified biomaterials and matrix surface. Blanchar, A. P. etal. (Biosensors and Bioelectronics, 1996, 11(6/7): 687-690) disclosescoating photoresists onto a substrate and then development using themicro-electromechanical mask to form on-spot hydrophilic spots, whereinthe region outside of the spots is hydrophobic. The preparation ofhigh-density nucleic acid probe microarrays and in situ synthesis iscarried out on this treated surface.

In these prior arts, blending, grafting or co-polymerization of ahydrophobic material and a compound bearing functional groups such asanhydride, imide, cyclic amide, and cyclic ester to prepare ahydrophobic copolymer is not disclosed. Further, the application of theprepared hydrophobic copolymer onto an organic or inorganic substrate toform an on-spot hydrophilic enhanced slide is also not disclosed.

SUMMARY OF THE INVENTION

It is therefore a primary object of the present invention to provide amethod for preparing an on-spot hydrophilic enhanced slide, comprising:(a) preparing a hydrophobic copolymer in a solvent to obtain a solutionof hydrophobic copolymer; (b) coating said solution of hydrophobiccopolymer onto a substrate (e.g. organic or inorganic substrate); and(c) removing said solvent.

Another aspect of the present invention provides an on-spot hydrophilicenhanced slide, comprising: (i) a substrate (e.g. organic or inorganicsubstrate); and (ii) a layer of hydrophobic functional groups formed bya hydrophobic copolymer, wherein said layer is coated onto saidsubstrate.

Yet another aspect of the present invention provides a non-spothydrophilic enhanced microarray, comprising: (i) a substrate (e.g.organic or inorganic substrate); (ii) a layer of hydrophobic functionalgroups formed by a hydrophobic copolymer, wherein said layer is coatedonto said substrate; and (iii) a biologically active material, which isimmobilized onto said layer of hydrophobic functional groups.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention will be more fully understood and furtheradvantages will become apparent when reference is made to the followingdescription of the invention and the accompanying drawings in which:

FIG. 1 is a schematic diagram showing the synthesis of PSMI.

FIG. 2 is a comparative diagram showing the fluorescence intensity afterimmobilization of labeled oligonucleotide probe using a glass slidecoated with PS and PSMI, respectively as a matrix.

FIG. 3 is a comparative diagram showing the fluorescence intensity afterimmobilization of labeled oligonucleotide probe using a glass slidecoated with PS and PSMI, respectively as a matrix.

FIG. 4 is a diagram showing changes of contact angle by acid/basesolution on the PSMA matrix of the present invention.

FIG. 5 is a diagram showing in-situ changes of contact angle by water,buffer, or oligonucleotide probe on the PSMA slide of the presentinvention.

FIG. 6 is a diagram showing the immobilization efficiency of the PSMAmatrix of the present invention.

FIG. 7 is a diagram showing the effect of ionic strength and pH on theimmobilization of oligonucleotide probe.

FIG. 8 is a diagram showing the fluorescence intensity afterhybridization.

FIG. 9 is a diagram showing the fluorescence intensity using mCOCsubstrate coated with PSMA as a matrix.

FIG. 10 is a diagram showing glass substrate coated with various ratiosof PSMA, wherein FIG. 10 a shows the fluorescence intensity afterimmobilization of oligonucleotide probe Sp5; and FIG. 10 b shows changesof contact angle by water.

FIG. 11 is a diagram showing a glass substrate coated with PSMA, whereinFIG. 11 a shows the fluorescence intensity after immobilization ofoligonucleotide probe Sp5; and FIG. 11 b shows the relative standarddeviation (R.S.D.) of the immobilization efficiency.

FIG. 12 is a diagram showing the fluorescence intensity using aglass-based matrix coated with PEMA.

DETAILED DESCRIPTION OF THE INVENTION

The present invention features a method for preparing an on-spothydrophilic enhanced slide by (a) preparing a hydrophobic copolymer in asolvent to obtain a solution of hydrophobic copolymer; (b) coating thesolution of hydrophobic copolymer onto a substrate; and (c) removing thesolvent. In accordance with the method of the present invention, thehydrophobic copolymer is prepared by blending, grafting orco-polymerization of a hydrophobic material and a compound bearingfunctional groups (e.g. anhydride, imide, cyclic amide, or cyclicester). The resulting hydrophobic copolymer is then coated onto anorganic or inorganic substrate to form a hydrophobic layer with covalentbonding functional groups, which is useful in the preparation of ahigh-density microarray. In addition, the covalent bonding reaction isreduced to one-step reaction so that the immobilization efficiency canthus be improved. Further, the compound bearing functional groups (e.g.anhydride, imide, cyclic amide, and cyclic ester) or the derivativesthereof, can be blended, grafted, or co-polymerized with a hydrophobicmaterial in various ratios to adjust the density of functional groups onthe matrix surface which can be provided for covalent bonding. Moreover,another hydrophilic group will be formed when the structure ofanhydride, imide, cyclic amide, or cyclic ester is attacked by abiomaterial or a modified nucleophile, which leads to the formation ofon-spot hydrophilic enhancement and thus bears positive or negativecharge. It is therefore beneficial to enhance not only the covalentbonding on the microarray but also the specificity of subsequentlybiochemical reaction, thereby improving the choke point of thetraditional hydrophilic silanization.

The term “on-spot hydrophilic enhanced” used herein refers to theformation of another hydrophilic group via ring-opening when thestructure of anhydride, imide, cyclic amide, or cyclic ester on thehydrophobic matrices prepared by the present invention is attacked by anucleophile (e.g. amine modified oligonucleotide probe), which leads tothe formation of on-spot hydrophilic enhancement. At this point, thehydrophobic surface is converted to hydrophilic, whereas the otherregion remains hydrophobic.

Another advantage of the present invention is that the negative charge(for example, anhydride group) or positive charge (for example, imide,cyclic amide) can be formed at the on-spot hydrophilic enhanced regionunder an appropriate condition when the ring-opening reaction occurs.This property improves the orientation of the immobilized biomaterials.

The schemes of the aforementioned reaction are shown below:

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. In addition, the abbreviationsthroughout the specification have the following meanings: PS,polystyrene; MA, maleic anhydride; PSMA, poly(styrene-co-maleicanhydride); MI, maleimide; PSMI, poly(styrene-co-maleimide); PE,polyethylene; and PEMA, poly(ethylene-co-maleic anhydride).

According to the method for preparing an on-spot hydrophilic enhancedslide of the invention, the type of the substrate used herein is notlimited, and can include an organic or inorganic substrate (i.e.substrate-independent). Organic substrates include a polymer polymerizedby organic monomers. Suitable organic monomers include, for example,ethylene, styrene, propylene, ester, acrylic acid, acrylate, alkylacrylic acid, or alkyl acrylate. Inorganic substrates include, but arenot limited to silicon wafer, ceramic material, glass or metal. In onepreferred embodiment, the hydrophobic copolymer prepared by the presentinvention (as described below) can be directly molded by injection orcompression to form an on-spot hydrophilic enhanced slide withhydrophobic surface, wherein the technique of injection or compressionmolding is well known to those skilled in this art. Suitable hydrophobiccopolymers used herein include, but are not limited to,poly(styrene-co-maleic anhydride), poly(styrene-co-maleimide), orpoly(ethylene-co-maleic anhydride).

If an inorganic substrate is employed, an activation step of thesubstrate surface can be carried out prior to coating hydrophobiccopolymers thereon to enhance the adhesion between the substrate surfaceand hydrophobic copolymers. The activation step includes treatment ofthe substrate surface with an acid or a base, or treatment of thesurface by plasma activation.

According to the preparation method of the invention, the substratesurface can be cleaned prior to coating hydrophobic copolymers thereonto prevent the deposition of impurities or contaminants on the substratesurface. The cleaning step is performed by pretreatment with a solventand/or sonication, based on the material of the substrate. Suitablesolvents include, but are not limited to surfactant, water, alcohol oracetone.

In accordance with the present invention, the hydrophobic copolymer isprepared by blending, grafting or co-polymerization of a hydrophobicmaterial and a compound bearing a functional group such as anhydride,imide, cyclic amide, or cyclic ester. The hydrophobic material caninclude any compound that blends, grafts or co-polymerizes with ananhydride, such as maleic anhydride, 2-decenylsuccinic anhydride,hex-2-enylsuccinic anhydride, tetrahydrophthalic anhydride; imide, suchas maleimide, 4-amino-phthalimide; cyclic amide, such as3-methyl-2-pyridone; or cyclic ester, such as 5-methylfuranone,4-aminophthalide, tetronic acid. Such hydrophobic materials includestyrene, urethane, ethylene, or derivatives thereof. In a particularlypreferred embodiment of the present invention, the hydrophobic materialused is polystyrene or ethylene, and the prepared hydrophobic copolymersthus include PSMA, PSMI, or PEMA.

The method for coating the hydrophobic copolymer onto a matrix is notlimited, and is understood by one of ordinary skill in the art to whichchemical engineering and semiconductor process belongs, to include spincoating, screen printing, roller coating, curtain coating, or dipcoating, etc. In one preferred embodiment, the coating method usedherein is spin coating, preferably at 3,000-6,000 rpm, and morepreferably at 4,000 rpm.

After coating the hydrophobic copolymer onto a substrate, the excesssolvent is removed by means of, for example, vacuum evaporation, heatingevaporation, or evaporation under reduced pressure, wherein the methodof heating evaporation is carried out at a temperature not higher than100° C. to prevent the matrix from being destroyed or to preventundesired polymerization. The preparation of the slide of the inventionis accompanied after the solvent is removed.

In another aspect of the present invention, an on-spot hydrophilicenhanced microarray is provided, comprising a biologically activematerial, which is immobilized onto the matrix described above. Theimmobilization is achieved by way of contacting the bio-molecules withthe on-spot hydrophilic enhanced slide of the present invention. Thelinkage between a nucleophile such as an amine group, which is presentin the molecule (e.g. protein) itself or in a chemically-modifiedentity, and functional groups such as anhydride, imide, cyclic amide orcyclic ester on the matrix surface is formed via the ring-openingreaction, whereas the other side of the ring bears negative charge (e.g.anhydride group) or positive charge (e.g. imide, cyclic amide), therebyimproving the orientation of the immobilized biomaterials.

Bio-molecules used as the biologically active material that are suitablefor use in the invention include nucleic acid, oligonucleotide, peptidenucleic acid (PNA), antigen, antibody, enzyme, or protein. Stable amidelinkages are formed after such bio-molecules are reacted with thefunctional groups of the on-spot hydrophilic enhanced slide of theinvention. As compared with the prior arts in which the bonding iscreated via a two-step reaction (i.e., by the silane-based polymer andfollowed by adding a crosslinker such as glutaldehyde), the reaction isreduced to one-step reaction in the present invention. Thereby, the timefor the immobilization reaction is substantially decreased andefficiency is increased.

Another feature of the present invention is the property of on-spothydrophobic/hydrophilic dynamic conversion when the slide is applied toimmobilization of the biomaterials. In addition, a high-densitymicroarray can be prepared according to the matrix with a functionallayer of hydrophobic characteristic, and the biomaterials' orientationduring subsequent reaction can be improved by way of the on-spothydrophilic conversion. Further, the preparation time according to themethod of the invention is much less than that of the conventionalmethod, and the homogeneity of the slide of the present invention isincreased. In other words, the number of the functional groups (e.g.anhydride, imide, cyclic amide, or cyclic ester) appearing on the matrixand the bonding strength is average. Moreover, the on-spot hydrophilicenhanced microarray of the present invention can markedly decrease thetime required for immobilization of biomaterials onto the matrix. In onepreferred embodiment, the immobilization time is less than 40 minutes.

Without intending to limit it in any manner, the present invention willbe further illustrated by the following examples.

EXAMPLE 1 Preparation of PSMI

The preparation scheme is shown in FIG. 1. To a pre-filled nitrogen orargon reaction bottle containing toluene, polystyrene and maleimide wereadded in a molar ratio of 4:1, followed by addition of 0.5%N,N′-azobisisobutyronitrile dropwise. The co-polymerization wasconducted at 60° C. for 6 hours, and then ceased by aeration to obtainPSMI in toluene.

EXAMPLE 2 Effect of PSMA and PS on Immobilization of Oligonucleotide

PSMA (Aldrich, Cat. No. 44240-2) and PS (Aldrich, Cat. No. 43010-2) weredissolved in toluene, respectively, and then coated onto glasssubstrates at 4,000 rpm to form PSMA and PS slides. The slides weredried in an oven at 100° C. to remove toluene. A syntheticoligonucleotide probe Sp5 composed of 25 nucleotides in which the 5′ endwas labeled with fluorescence and the 3′ end bore amine group, wasimmobilized to the aforementioned slides to perform a ring-openingreaction. The immobilization conditions were described as follows: 0.5μM of Sp5 in 2×SSC buffer (pH 7.0) was spotted on the slides andincubated at 37° C. for 1 hour. The slides were washed with 0.2% SDS for10 minutes. The fluorescence analyses were performed for the slides withand without washing (control) to monitor the immobilization efficiency.The result is shown in FIG. 2.

EXAMPLE 3 Effect of PSMI and PS on Immobilization of Oligonucleotide

PSMI synthesized from Example 1 and PS were coated onto glasssubstrates, respectively, according to the method described in Example2. The slides were dried in the oven at 100° C. to remove solvent. Asynthetic oligonucleotide probe Sp5 as set forth above was immobilizedto the slides to perform a ring-opening reaction. The immobilizationconditions were similar to those in Example 2. The fluorescence analyseswere performed for the slides with and without washing (control) tomonitor the immobilization efficiency. The result is shown in FIG. 3.

EXAMPLE 4 Effect of Acid/Base Treatment on the Surface Hydrophilicity

PSMA slides prepared from Example 2 were spotted with 1 N HCl and 1 NNaOH, respectively. The contact angle between the droplet and slide wasmeasured at 0, 10, 20, and 30 minutes. The result is shown in FIG. 4.

EXAMPLE 5 Effect of Oligonucleotide Immobilization on the SurfaceHydrophilicity

PSMA slides prepared from Example 2 were spotted with de-ionized water,Sp5 probe (pH 7; as set forth above), and buffer (pH 7). The contactangle between the droplet and slide was measured at 0, 10, 20, and 30minutes. The result is shown in FIG. 5.

EXAMPLE 6 Effect of Various Parameters on Immobilization ofOligonucleotide

PSMA slides were used in this example to test the effect of parametersincluding time, ionic strength, and pH on the immobilization ofoligonucleotide. The immobilization conditions were similar to those inExample 2, wherein the probes used were Sp5 (as set forth above) and O₃(without amine modification at the 5′ end). The fluorescence wasmeasured at 10, 30, and 60 minutes to evaluate the relationship ofimmobilization efficiency with time. In addition, probes were dissolvedin 0, 0.1, 0.5, 1.0, 2.0, 4.0, and 6.0 M NaCl solution forimmobilization, respectively, to evaluate the relationship betweenimmobilization efficiency and ionic strength. Further, probes wereimmobilized under the environment of pH 3, pH 7, and pH 11 to evaluatethe optimal conditions for immobilization. The results are shown in FIG.6 and 7.

EXAMPLE 7 Hybridization with Specific Oligonucleotides

The slides used in this Example were the same as Example 2. Theoligonucleotides used in this example were AO₃ (composed of 29nucleotides with amine group at the 5′ end); O₃ (composed of 29nucleotides without amine group at the 5′ end); and ATO₃ (composed of 15thymidine bases and the same 29 nucleotides as set forth above withamine group at the 5′ end), respectively. The probe used for labelinghybridization reaction was the complementary sequence thereto, whereinthe 3′ end was labeled with fluorescence. The immobilization conditionswere 2×SSC, pH 7.0; 3×SSC, pH 7.0; and 10×SSC, pH 3.0, respectively, for1 hour. The hybridization reaction was performed for 4 hours. Thefluorescence was analyzed to monitor the hybridization result. Theresult is shown in FIG. 8.

EXAMPLE 8 Effect of Organic Slide

PSMA was coated onto metallocene cycloolefine copolymers (mCOC)substrates at 4,000 rpm. The slides were dried in the oven at 100° C. toremove solvent. A synthetic oligonucleotide probe Sp5 as set forth abovewas immobilized to the aforementioned slides to perform a ring-openingreaction. The immobilization conditions were similar to those in Example2. The fluorescence analyses were performed for the slides with andwithout washing (control) to monitor the immobilization efficiency. Theresult is shown in FIG. 9.

EXAMPLE 9 Effect of Various Proportions of PSMA on Immobilization ofOligonucleotide

PSMA in the range of 2%-44% was coated onto glass substrates,respectively. The coating method and conditions were the same as Example8 to evaluate the immobilization efficiency of oligonucleotides. Theresult is shown in FIG. 10 a. In addition, de-ionized water was spottedonto the matrices and the contact angle between the droplet and surfaceof matrix was measured at various composition of PSMA matrices. Theresult is shown in FIG. 10 b.

EXAMPLE 10 RSD Test

PSMA slides prepared from Example 2 were used for immobilization of Sp5probe for 60 minutes under the same conditions. The fluorescence wasanalyzed for each slide, followed by calculation of the relativestandard deviation (R.S.D.) of the immobilization efficiency. The resultis shown in FIG. 11.

EXAMPLE 11 Effect of PEMA on Immobilization of Oligonucleotide

PEMA (Aldrich, Cat. No. 43084-6) was coated onto glass substratesaccording to the method described in Example 2. The slides were dried inthe oven at 100° C. to remove solvent. A synthetic oligonucleotide probeSp5 as set forth above was immobilized to the slides to perform aring-opening reaction. The immobilization conditions were similar tothose in Example 2. The fluorescence analyses were performed for theslides with and without washing (control) to monitor the immobilizationefficiency. The result is shown in FIG. 12.

The PSMA, PSMI, and PEMA hydrophobic copolymers of the present inventionpersist reaction functionalities (e.g. anhydride ring or imide ring) towhich the biomaterials can be bonded, so that a bio-molecule such asamine-modified DNA can be stably bonded to the surface of the slides.Referring to FIGS. 2-3, because PS itself cannot provide any functionalsite for bonding, the oligonucleotide probes were washed out afterspotting Sp5 and washing with 0.2% SDS, thus no fluorescent signal canbe detected (i.e. biomaterials are not stably adhered to the surfaces).As compared with PSMA or PSMI slides, the Sp5 probe retains at least 90%immobilization efficiency after washing with 0.2% SDS.

Referring to FIG. 4, the contact angle is reduced from 120° to 90° byacid/base treatment on PSMA slide. By contrast, referring to FIG. 5, thecontact angle is not reduced markedly by buffer solution(pH 7) andde-ionized water on the slide, indicating there is no hydrophilicconversion occurred thereon, i.e. no ring-opening reaction occurred.After spotting Sp5 probe (pH 7) on the slide, the contact angle isreduced from 120° to 90°, showing the nucleophile (amine) attacks theanhydride ring. This results in ring-opening reaction and bondingbetween the DNA and slide. Meanwhile, the hydrophilicity is enhanced.

In the conventional method, about 4-16 hours are required for theimmobilization of oligonucleotide probe. As shown in FIG. 6, it takesonly about 30 minutes for the same effect on the on-spot hydrophilicenhanced slide of the present invention, which is significantly shorterthan the prior technique.

Referring to FIG. 7, under the conditions of 0 to 0.1 M ionic strengthand pH 7 to 11, the preferred immobilization efficiency is obtained onthe on-spot hydrophilic enhanced slide of the present invention.

Referring to FIG. 8, the differences among three probes are that AO₃bears an amine group at the 5′ end, whereas O₃ is without an aminegroup. The amine group at the 5′ end of ATO₃ is ligated with 15 bases ofthymidine. Three kinds of probe are immobilized onto the slide at pH 3.0and 7.0, respectively. From the result of O₃ probe, the poor bondingefficiency is observed due to the lack of an amine group at the 5′ end.Other probes with amine group at the 5′ end possess excellent bondingefficiency. This indicates the specificity of the bonding of the targetmolecule can be elevated markedly according to the on-spot hydrophilicenhanced slide of the present invention.

FIG. 9 shows the matrix used in the invention is not limited toinorganic substrate. The metallocene cycloolefine copolymer (mCOC) is anorganic polymer, and the hydrophobic copolymer of the present inventioncan be still coated thereon to form a thin layer, which is useful forthe subsequent immobilization of biomaterials.

The property of hydrophobicity differs from the proportions of thehydrophobic copolymers, and thus the density of functional groupsprovided for immobilization is also different. Referring to FIG. 10 a,the higher anhydride content, the more hydrophilic is obtained. However,the main chain of the copolymer remains hydrophobic, which is beneficialfor the preparation of high-density slides. Meanwhile, referring to FIG.10 b, it is shown that the higher anhydride content, the greaterimmobilization efficiency obtained. However, the higher hydrophilicityis not suitable for preparing high-density slides. The hydrophobiccopolymer containing too much anhydride content is therefore notsuitable.

Homogeneity is important for the preparation of microarrays. Referringto FIG. 11, the homogeneity of the hydrophobic copolymer is excellentafter coating to the substrate. It shows excellent immobilizationefficiency and less than 2% relative standard deviation (R.S.D.).

The component of the hydrophobic copolymer is not limited tostyrene/polystyrene. As shown in FIG. 12, the polyethylene (PE) used asthe main chain of the copolymer can also develop into an on-spothydrophilic enhanced slide. Any functionally hydrophobic copolymercontaining at least one anhydride, imide, cyclic amide, or cyclic estercan attain the purpose of the present invention.

While the invention has been particularly shown and described with thereference to the preferred embodiment thereof, it will be understood bythose skilled in the art that various changes in form and details may bemade without departing from the spirit and scope of the invention.

1. An on-spot hydrophilic enhanced slide, comprising: (i) a substrate,said substrate comprising an organic substrate or an inorganicsubstrate, wherein said organic substrate comprises a polymerpolymerized by organic monomers, wherein said organic monomers areselected from the group consisting of ethylene, styrene, propylene,ester, acrylic acid, acrylate, alkyl acrylic acid, and alkyl acrylate,wherein said inorganic substrate comprises silicon wafer, ceramicmaterial, glass or metal; and (ii) a functionally active layer formed bya hydrophobic copolymer, wherein said hydrophobic copolymer comprises ahydrophobic material and a compound bearing a functional group, whereinsaid functional group of said compound is at least one selected from agroup consisting of of anhydride, imide, cyclic imide, and cyclic ester,wherein said layer is coated onto said substrate.
 2. The slide asclaimed in claim 1, wherein the hydrophobic copolymer is prepared byblending, grafting or co-polymerization of the hydrophobic material andthe compound bearing the functional group.
 3. The slide as claimed inclaim 2, wherein the hydrophobic material is selected from the groupconsisting of styrene, urethane, ethylene, and derivatives thereof. 4.The slide as claimed in claim 3, wherein the hydrophobic copolymercomprises poly(styrene-co-maleic anhydride), poly(styrene-co-maleimide),or poly(ethylene-co-maleic anhydride).
 5. The slide as claimed in claim1, wherein the organic substrate comprises poly(styrene-co-maleicanhydride), poly(styrene-co-maleimide), or poly(ethylene-co-maleicanhydride).